Inhibition of c-Src blocks oestrogen-induced apoptosis and restores oestrogen-stimulated growth in long-term oestrogen-deprived breast cancer cells

Abstract Purpose Our publications demonstrate that physiological concentrations of oestrogen (E 2 ) induce endoplasmic reticulum and oxidative stress which finally result in apoptosis in E 2 -deprived breast cancer cells, MCF-7:5C. c-Src is involved in the process of E 2 -induced stress. To mimic the clinical administration of c-Src inhibitors, we treated cells with either E 2 , a c-Src inhibitor PP2, or the combination for 8 weeks to further explore the apoptotic potential of the c-Src inhibitor and E 2 on MCF-7:5C cells. Methods Protein levels of receptors and signalling pathways were examined by immunoblotting. Expression of mRNA was detected through real-time polymerase chain reaction (PCR). Cell cycles were analysed by flow cytometry. Results Long-term treatment with PP2 alone or E 2 alone decreased cell growth. In contrast, a combination of PP2 and E 2 blocked apoptosis and the resulting cell line (MCF-7:PF) was unique, as they grew vigorously in culture with physiological levels of E 2 , which could be blocked by the pure antioestrogen ICI182,780. One major change was that PP2 collaborated with E 2 to increase the level of insulin-like growth factor-1 receptor beta (IGF-1Rβ). Blockade of IGF-1Rβ completely abolished E 2 -stimulated growth in MCF-7:PF cells. Furthermore, combination treatment up-regulated transcription factors, Twist1 and Snail, and repressed E-cadherin expression which made MCF-7:PF cells display a characteristic phenotype of epithelial–mesenchymal transition (EMT). Conclusions These data illustrate the role of the c-Src inhibitor to block E 2 -induced apoptosis and enhance E 2 -stimulated growth. Caution must be exercised when considering c-Src inhibitors in clinical trials following the development of acquired resistance to aromatase inhibitors, especially in the presence of the patient’s own oestrogen.